PROJECT SUMMARY DNA Damage Response (DDR) pathways, including DNA repair and cell-cycle checkpoints, are critical for protecting against genomic insults and thus for maintaining genome integrity. The most deleterious type of DNA damage is the DNA double-strand break (DSB), and deficiencies in the DDR DSB subset repair pathways result in genomic instability via chromosomal aberrations and mutations, which can ultimately lead to an increased risk for disease, including cancer. However, the mechanisms regulating DSB repair pathways have yet to be fully elucidated, leaving us with an incomplete understanding of how dysregulation or mutation of proteins involved in DSB repair pathways increases the risk for developing cancer. Our lab has recently identified Sirtuin 2 (SIRT2), a deacetylase and known murine tumor suppressor, as a novel regulator of the DDR. SIRT2-deficient cells are hypersensitive to DNA damage and show impaired checkpoint responses following ionizing radiation. Preliminary data indicate that SIRT2 is necessary for the proper repair of DSBs through the homologous recombination (HR) repair pathway and is a novel regulator of Breast Cancer 1 (BRCA1), a tumor suppressor and critical regulator of HR. Our findings suggest that SIRT2 is an important player in the regulation of DSB repair and may contribute to the development of cancers that arise from faulty DSB repair pathways. However, the precise mechanism that SIRT2 plays in the repair of DSBs remains unclear. In this project, a combination of genetic, biochemical, molecular biological, and cell biological approaches will be used to test the hypothesis that least in part, by promoting BRCA1 function in HR repair through deacetylation. The specific aims of this research proposal are 1) to determine the functional significance of the SIRT2/BRCA1 interaction during HR repair and 2) to determine the sites of SIRT2 deacetylation on BRCA1 in response to DNA damage and their functional significance. Insights gained from completion of this work will contribute to our fundamental knowledge of DSB repair pathways and how dysregulation of DSB repair by the SIRT2 and BRCA1 tumor suppressor proteins leads to genomic instability and an increased risk for cancer. Additionally, these findings will provide invaluable insights into how to exploit the interplay between SIRT2 and BRCA1 as a novel therapeutic approach for the prevention and treatment of cancer. SIRT2 maintains genome integrity in response to DSBs, at